The long-term objective of this project has been to develop Tetrahymena thermophila as a useful microbial eukaryotic model system for the study of any mechanism of interest in cell and molecular biology. Our approach exploits the utility and versatility of conditional lethal mutations. We no propose 1) to build a linkage map of chromosome 2 by deletion mapping (our share of a wider Tetrahymena mapping endeavor); 2) to identify nonsense mutations and suppressors and to develop an efficient method for quickly identifying new mutations as nonsense; 3) to isolate mutants blocked at 2 key fertilization events (reciprocal exchange and fusion of pronuclei, respectively) and 4) to complete on-going studies: characterizing vinblastine resistant mutants and studying the onset of gene expression in the new differentiating macronucleus. In combination, these studies are significant in a variety of contexts. 1) They will increase the value of T. thermophila as a useful (and, in some aspects, unique) model system for the genetic dissection of cellular and molecular mechanisms. 2) They will open the way to isolate mutations blocking the profoundly significant mechanims of somatic DNA rearrangements. 3) They will increase our understanding of genetic mechanims operating in Tetrahymena, both during conjugation and in the macronucleus (differentiating and mature). 4) Concepts, strains and methods derived from these studies may find experimental or clinical application to human health-related problems, such as phenylketonuria (PKU), or immunoglobins and their pathologies.